Project Summary Inhibition of Wnt Receptor Activation by the Tumor Suppressor Adenomatous Polyposis Coli The long-term objective of this study is to investigate how the tumor suppressor Adenomatous polyposis coli (APC) inhibits the Wnt signal transduction pathway by regulating the Wnt receptor complex (signalosome) and to demonstrate how this can be exploited to target APC mutant colorectal cancers (CRCs). Wnt signaling is essential for intestinal stem cell maintenance, whereas aberrant activation of this pathway, which occurs most frequently through mutational inactivation of APC, triggers the development of the vast majority of CRCs. In the classical model for Wnt signaling, the sole role of APC is to destabilize the key transcriptional activator in the Wnt pathway, beta-catenin. However, our recently published findings reveal an additional and entirely new function ? APC prevents the internalization and consequent activation of the signalosome, a novel role that is evolutionarily conserved. We have shown that: 1) inducible loss of APC is rapidly followed by ligand-independent signalosome activation; 2) depletion or antibody-mediated inhibition of LRP6 (a signalosome component) inhibits the stabilization of beta-catenin, the transcriptional activation of Wnt target genes, and the proliferation of APC mutant cells; and 3) in APC mutant cells, endocytosis of Wnt receptors is required for the aberrant activation of Wnt signaling. The goal of this project is to use in vitro, ex vivo, and in vivo approaches to gain a better understanding of how APC inhibits signalosome activation under physiological conditions and to determine how aberrant activation of the signalosome underlies the consequences of APC inactivation in tumors. The three specific aims are to: 1) elucidate the mechanism by which APC loss promotes signalosome assembly in CRC cells; 2) identify the APC mutant CRC cells most susceptible to LRP6 inactivation; and 3) test the efficacy of LRP6 inactivation on CRC tumorigenicity in vivo. Because the molecular mechanisms by which APC prevents the aberrant activation of Wnt signaling are important for our understanding of colorectal carcinogenesis, the knowledge gained from this study will aid in the development of new therapeutic strategies for the treatment of CRC and other Wnt-driven cancers.